The Analysis of Jojoba Oil by LC-Coordination Ion Spray-MS (LC-CIS-MS)

Transcription

1 The Analysis of Jojoba Oil by LC-Coordination Ion Spray-MS (LC-CIS-MS) K. Lazou 1, A. d Oosterlinck 2, L. De Reu 1, M. Schelfaut 2, and P. Sandra 2 * 1 Department of Chemical Quality Control, Janssen Pharmaceutica, B-244 Geel, Belgium 2 Department of Organic Chemistry, University of Gent, Krijgslaan 281 S4, B-9 Gent, Belgium, pat.sandra@rug.ac.be Key Words: microbore LC; coordination ion spray-mass spectrometry (CIS-MS); jojoba oil; wax ester composition Summary Liquid chromatography combined with coordination ion spray mass spectroscopy (LC-CIC- MS) was evaluated for the analysis of the wax esters of jojoba oil. Excellent ionisation was obtained to elucidate the molecular ion [M+Ag] + of the wax esters and, moreover, by applying collision induced dissociation (CID) at 29 V both the acidic and alcoholic part could be identified. 1

2 1 Introduction Jojoba oil is obtained from the seeds of the evergreen shrub, Simmondsia chinensis, which is native to the Sonora Desert (U.S.). Because of its economic value, the plant is now cultivated in many arid and semiarid countries all over the world. The seeds contain about 5% of a light yellow, odorless oil commonly referred to as jojoba oil and consisting of long-chain esters rather than a mixture of triglycerides, which gives the Jojoba oil its unique characteristics [1]. The oil is extensively used in the cosmetic industry for its dermatological properties. It is actually also applied as high pressure lubrificant and as plasticizer for polymers. Jojoba oil may be considered as a low-energy replacement for conventional fats and oils in diet products as it is poorly digested. Analysis of jojoba oil is normally performed after hydrolysis. The acids, converted to the methyl esters and the alcohols are analysed by GC [1,2]. This contribution describes the analysis of the wax esters of jojoba oil as such by Liquid Chromatography-Coordination Ion Spray-Mass Spectrometry (LC-CIS-MS). 2 Experimental 2.1 Instrumentation LC-MS was carried out on a benchtop HP11 series LC/MS (Hewlett-Packard, Waldbronn, Germany) equipped with in-source collision induced dissociation (CID) to obtain additional fragmentation. A syringe pump, type apparatus 22 (Harvard Apparatus, Massachusetts, USA), was used to pump a AgNO 3 solution from a 5 ml syringe (SGE, Austin Texas, USA) into the mobile phase flow via a post column T-piece (Valco, Houston, USA). 2.2 Chemicals All solvents were HPLC grade from Labscan (Dublin, Ireland) and AgNO 3 was analytical grade from Aldrich (Bornem, Belgium). Jojoba oil was obtained by cold pressing jojoba seeds. The standards of cis-9-octadecenoyl - cis-9-octadecenoate, decanoyl - cis-9- octadecenoate and hexyl-dodecanoate were purchased from Henkel (Dusseldorf, Germany). 2

3 2.3 Analytical Conditions The column used was a Spherisorb 25 long x 2. mm I.D., 5 µm, BDS from Waters (Milford, USA). The mobile phase consisted of MeOH/acetone/hexane (2/1/1). The flow rate was. ml/min. The AgNO 3 solution had a concentration of 1.6 mg/ml and the post column addition flow was 1 µl/min. The injection volume was.5 µl of a 1 mg/ml hexane solution. The AP-ESI interface was set in the positive ionisation mode with following parameters: drying gas temperature: C; drying gas flow: 12 L/min; nebulizing gas flow: 45 psig; capillary voltage: 5 V; mass scan range: 1-8 m/z; fragmentor voltage at 1 V or 29 V. 3 Results and Discussion Although AP-CI and, in less extent AP-ESI, can be applied for the structure elucidation of triglycerides [3], both failed completely for the analysis of the wax esters of jojoba oil. Very noisy baselines and nearly no ionization were observed. Also other classes of compounds such as terpenes, sugars, alcohols, aromatic compounds and vitamines gives, in both AP-CI and AP-ESI, very poor detectability. The sensitivity for those compounds can be increased by modifying the mobile phase composition. To keep the separation profile unaffected postcolumn addition can be applied. Karlsson [4] dsecribed cationization of the analytes with alkali metals (sodium, potassium, rubidium and cesium). By using concentrations of 5 x 1-5 M of the acetate form of each metal, the detection limits for cyclodextrins, oligosaccharides and bafilomycins were significantly increased. Kohler et al. [5] experimented with post column addition of a metal chloride solution for the analysis of carbohydrates. Unlike postcolumn derivatization the process of complex formation is immediate and no post column reactor is needed. Further investigations by E. Bayer et al. [6,7] led to a new method of chemical ionization in which positively or negatively charged complexes are formed by the addition of a suitable atom to the analytes, followed by detection of the formed complexes by mass spectrometry. Since both polar and nonpolar organic compounds can form coordination compounds with an appropriate central atom, this form of ionization is highly versatile. This new technique is referred to as Coordination Ion Spray - Mass Spectrometry (CIS-MS). Neither an electric field nor the formation of a reagent gas plasma by corona discharge is needed for ionization. Only, efficient nebulization in the ion source is mandatory. Moreover, the sample is subjected to less thermal stress than when using AP-CI. 3

4 Olefins and polyolefins such as terpenes, sesquiterpenes, carotenoids vitamines and steroids are particularly attractive solutes to consider CIS-MS. With metals of the first transition groups, for example Cu I, Ni II, Pd II, Pt, and Ag I, these unsaturated compounds form highly stable π-complexes. Before LC-CID-MS of jojoba oil with post-column silver ion addition, some standard solutions were introduced with flow injection analysis (FIA) with the T-piece setup. The solutes comprised an unsaturated ol-unsaturated acid ester (cis-9-octadecenoyl - cis-9- octadecenoate) an saturated ol-unsaturated acid ester (decanoyl - cis-9-octadecenoate) and a saturated ol-saturated acid ester (hexyl-dodecanoate). The spectra are shown in Figure 1. Pseudo molecular ion signals ([M+Ag] + ) are clairly observed and with excellent detectability. A B C D Figure 1: CIS-MS analysis of standard samples. A: cis-9-octadecenoyl - cis-9-octadecenoate, fragmentor voltage 1 V; B: octadecenoyl - cis-9-octadecanoate, fragmentor voltage 29 V, C: decanoyl - cis-9-octadecanoate, fragmentor voltage 29 V, D: hexyl-dodecanoate, fragmentor voltage29 V. 4

5 The Ag atom gives rize to isotopic peaks at m/z 2 with similar intensities. This means an extremely easy indication of any present Ag complex. Surprisingly, the saturated species was also complexed by silver? This phenomenon is presently under study and it seems that complexation also occurs at the carbonyl ester function. Increasing the fragmentor voltage causes, next to the pseudomolecular ions, fragment ions when the fatty acid or alcohol moieties have an unsaturated bond. Unsaturation allows an Agcomplex to be formed which introduces a positive charge and the fragment becomes detectable with good sensitivity (Figure 1 B & C). The observed m/z signal for the acid fragment part appears to be the MW of the entire acid molecule plus an Ag atom. Table 1: MW and m/z data of some long chain ester standards and their potential fragments. Ester compound Acid fragment Alcohol fragment C nr MW m/z MW m/z symbol MW m/z symbol Cis-9-octadecenoyl - Cis-9-octadecenoate C : :1 Decanoyl - Cis-9-octadecanoate C : hexyl-dodecanoate C m/z: mono-isotopic signal as detected with MS The alcoholic part is detected as a complex of silver minus the loss of water. If the acid or the alcohol part is saturated, no signals for that particular fragment are seen. In figure 1 C, the saturated decanoyl fragment remains undetected. For completely saturated esters no signal of neither of both fragments could be observed (Figure 1.D). The LC-CIS-MS analysis of jojoba oil is shown in Figure 2. 5

6 Figure 2: LC-CIS-MS analysis of jojoba oil The identification results are summarized in Table 2 where the actual detected signals in CIS-MS (m/z) are given next to the molecular weight (MW) of the entire esters and of each fragment. The retention time is mentioned together with the area % data as calculated using ion extraction. The total carbon number (CN) symbols for the fragments are also given. Table 2: Identification of jojoba oil constituents. Ester compound Acid fragment Alcohol fragment CN t R (min) area % MW m/z MW m/z symbol MW m/z symbol C : :1 C : :1 C : :1 C : :1 C : :1 C : :1 C : :1 C : : m/z: mono-isotopic signal as detected with MS In accordance with the literature [7], four major carbon numbers are identified, namely C 38, C 4, C 42 and C 44. Peak 2 is a composite of C18:1-C:1 (CN 38) with C17:-C19:1 (CN 36) and CN42 (peak 4) of C:1-C22:1 and C22:1-C:1. Quantification could be done via ion extraction of the specific fragments taking a response of 1 into account. To illustrate the quality of the spectra in LC-CID-MS, Figure 3 shows the obtained spectra at 29 V (CID) for peak 4 (major) and peak 6 (minor). 6

7 A m/z 1 B Figure 3: LC-CIS-MS of peaks 4 and 6 at 29 V CID. m/z 7

8 4 Conclusion LC-CIS-MS is a very easy and straightforward technique for the analysis of solutes which gives very poor ionization in AP-CI and AP-ESI. This was illustrated with the analysis of the wax esters of jojoba oil. References [1] M.L. Tonnet, R.L. Dunstone, JAOCS, 1984, 61, [2] M. Pina, D. Pioch, J. Graille, Lipids, 1987, 22, [3] P. Sandra, Y. Zhao, K. Lazou, A. Medvedovici, publication in preparation [4] K.-E. Karlsson, J. Chromatogr. A, 1998, 794, [5] M. Kohler, J. A. Leary, Anal. Chem., 1995, 67, [6] E. Bayer, P. Gfrörer, C. Rentel, Angew. Chem. Int. Ed., 1999, 38, [7] C. Rentel, P. Gfrörer, E. Bayer, Electrophoresis, 1999,,